`United States Patent
`[11] Patent Number:
`5,194,447
`
`Kao [45] Date of Patent: Mar. 16, 1993
`
`||||||lllllllllllllllllllllIllllllllllllll|||||llllllllllllllllllllllllllll
`USOOSl94447A
`-
`
`[54] SULFONYLCARBAMATES OF RAPAMYCIN
`
`[57]
`
`ABSTRACT
`
`[75]
`
`Inventor: Wenling Kan, Chester, Pa.
`
`A compound of the structure
`
`[73] Assignee: American Home Products
`Corporation, New York, NY.
`
`[21] Appl. No.: 837,048
`
`[22] Filed:
`
`Feb. 18, 1992
`
`[51]
`
`Int. Cl.5 ............... .. 0071) 491/16; A61K 31/395;
`A61K 31/685; CO7F 7/04
`[52] US. Cl. ................................... .. 514/542; 514/546
`[58] Field of Search ................................ 540/452, 456
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`....................... 424/192
`3,992,992 12/1975 Sehgal et a1.
`.. 424/122
`3,993,749 11/1976 Sehgal et al.
`
`4,316,885 2/1982 Rakhit ......
`.. 424/122
`
`4,401,653
`8/1983 Eng ..........
`.. 424/124
`
`4,650,803
`3/1987 Stella et a1.
`.. 514/291
`..
`
`4,885,171 12/1989 Sehgal et a1.
`.. 424/122
`
`5,073,999
`1/1992 Warner et a1.
`424/122
`5,080,899
`1/1992 Sturm er al.
`.. 424/122
`
`5,091,389 2/1992 Ondeyka ct a].
`514/291
`.
`
`5,100,883
`3/1992 Schiehser
`514/183
`5,1m,889
`3/1992 Calne ................................. .. 514/291
`OTHER PUBLICATIONS
`
`U.S. Ser. No. 07/686,727 filed Apr. 17, 199l—Kao, Vo-
`gel, Musser.
`Vezina, C. J. Antibiot. 28, 721—726 (1975).
`Sehgal, S. N., J. Antibiot. 28, 727-732 (1975).
`Baker, 11., J. Antibiot. 31, 539~545 (1978).
`Martel, Q, Can. J. Physiol. Pharmacol. 55, 48 (1977).
`Staruch et 31., FASEB 3, 341] (1989).
`Dumont, F, FASEB 3,5256(1989).
`Calne et al., Lancet, 1183—1185 (1978).
`Morris, R. Med. Sci. Res. 17:877 (1989).
`
`Primary Examiner—Robert T. Bond
`Attorney. Agent, or Firm—Amold S. Milowsky
`
`
`
`independently, hydrogen or
`
`wherein
`R1 and R2 are each,
`—CONHS02—Ar; and
`Ar is phenyl, naphthyl, pyridyl, quinolyl, isoquinolyl,
`quinoxalyl, thienyl, thionaphthyl, furyl, benzofuryl,
`benzodioxyl, benzoxazolyl, benzoisoxazolyl, or ben-
`zodioxolyl; wherein the Ar group may be optionally
`mono—, di. or tri-substituted; with the provision that
`R1 and R2 are not hydrogen; or
`or a pharmaceutically acceptable salt thereof which by
`virtue of its immunosuppressive activity is useful
`in
`treating transplantation rejection, host vs. graft disease,
`autoimmune diseases and diseases of inflammation; by
`virtue of its antitumor activity is useful in treating solid
`tumors; and by virtue of its antifungal activity is useful
`in treating fungal infections.
`
`7 Claims, No Drawings
`
`Par Pharm., Inc.
`Exhibit 1122
`
`Case IPR201 6-01 479
`
`Par Pharm., Inc. v. Novartis AG
`
`Ex. 1 122-0001
`
`Ex. 1122-0001
`
`
`
`1
`
`5,194,447
`
`2
`
`wherein
`independently, hydrogen or
`R1 and R2 are each,
`—CONHSOz-Ar; and
`Ar is phenyl, naphthyl, pyridyl, quinolyl, isoquinolyl,
`quinoxalyl, thienyl, thionaphthyl, furyl, benzofu-
`ryl, benzodioxyl, benzoxazolyl, benzoisoxazolyl, or
`benzodioxolyl; wherein the Ar group may be op-
`tionally mono, di-, or tri-substituted with a group
`selected from alkyl of 1—6 carbon atoms, arylalkyl
`of 7-10 carbon atoms, alkoxy of 1-6 carbon atoms,
`cyano, halo, nitro, carbalkoxy of 2-7 carbon atoms,
`tritluoromethyl, amino, dialkylamino of 1-6 carbon
`atoms per alkyl group, alkylthio of 1—6 carbon
`atoms, ——SO3H, —PO3H, and —-C02H;
`with the proviso that R1 and R2 are not both hydrogen;
`or a pharmaceutically acceptable salt thereof when the
`Ar group contains a basic nitrogen or when the Ar
`group is substituted by dialkylamino of 1—6 carbon
`atoms per alkyl group, --S03H, —P03H, or C0211.
`The phamaceutically acceptable salts are those de—
`rived from such inorganic cations such as sodium, po-
`tassium, and the like; organic bases such as: mono-, di-,
`and trialkyl amines of 1-6 carbon atoms, per alkyl group
`and mono-, di-, and trihydroxyalkyl amines of 1—6 car-
`bon atoms per alkyl group, and the like; and organic and
`inorganic acids as: acetic, lactic, citric, tartaric, suc-
`cinic, maleic, malonic, gluconic hydrochloric, hydro-
`bromic, phosphoric, nitric, sulfuric, methanesulfonic,
`and similarly known acceptable acids and.
`Of these compounds, preferred members are those in
`which Ar is optionally mono-, di-, or tri- substituted
`phenyl. When Ar is substituted with arylalkyl of 7-10
`carbon atoms, it is preferred that aryl portion of the
`arylalkyl moiety be a phenyl group.
`The compounds of this invention carbamylated at the
`42-position or at both the 31- and 42-positions can be
`prepared by reacting rapamycin with an isocyanate
`having the general structure
`
`10
`
`15
`
`20
`
`25
`
`30
`
`35
`
`SULFONYLCARBAMATES OF RAPAMYCIN
`
`BACKGROUND OF THE INVENTION
`
`This invention relates to carbamates of rapamycin
`and a method for using them in the treatment of trans-
`plantation rejection, host vs. graft disease, autoimmune
`diseases, diseases of inflammation, solid tumors, and
`fungal infections.
`Rapamycin is a macrocyclic triene antibiotic pro-
`duced by Streptomyces hygroscopicus, which was found
`to have antifungal activity, particularly against Candida
`albicans, both in vitro and in vivo [C. Vezina et al., J.
`Antibiot. 28, 721 (1975); S. N. Sehgal et al., J. Antibiot.
`28, 727 (1975); H. A. Baker et al., J. Antibiot. 31. 539
`(1978); US. Pat. No. 3,929,992; and Us. Pat. No.
`3,993,749].
`Rapamycin alone (U-.S. Pat. No. 4,885,171) or in com—
`bination with picibanil (U.S. Pat. No. 4,401,653) has
`been shown to have antitumor activity. R. Martel et a1.
`[Can. J. Physiol. Pharmacol. 55, 48 (1977)] disclosed
`that rapamycin is effective in the experimental allergic
`encephalomyelitis model, a model for multiple sclerosis;
`in the adjuvant arthritis model, a model for rheumatoid
`arthritis; and effectively inhibited the formation of IgE-
`like antibodies.
`The irnmunosuppressive effects of rapamycin have
`been disclosed in FASEB 3, 3411 (1989). Cyclosporin A
`and FK-506, other macrocyclic molecules, also have
`been shown to be effective as immunosuppressive
`agents, therefore useful in preventing transplant rejec-
`tion [FASEB 3, 3411 (1989); FASEB 3, 5256 (1989); R.
`Y. Calne et al., Lancet 1183 (1978); and European Pa-
`tent Application 0401747 A2].
`Mono- and diacylated derivatives of rapamycin (es-
`terified at the 28 and 43 positions) have been showa to
`be useful as antifungal agents (US. Pat. No. 4,316,885)
`and used to make water soluble prodrugs of rapamycin
`(U.S. Pat. No. 4,650,803). Recently,
`the numbering
`convention for rapamycin has been changed; therefore
`according to Chemical Abstracts nomenclature,
`the
`esters described above would be at the 31- and 42-posi-
`tions.
`'
`
`DESCRIPTION OF THE INVENTION
`
`This invention provides derivatives of rapamycin
`which are useful as immunosuppressive, anti-inflamma-
`tory, antifungal, and antitumor agents having the struc-
`ture
`
`
`
`O=C_—_N—-502-—Ar
`
`4s
`
`55
`
`65
`
`under neutral conditions or in the presence of a base,
`such as pyridine.
`The 31—carbamylated compounds of this invention
`can be prepared by protecting the 42-alcohol of rapa-
`mycin with a protecting group, such as with a tert-butyl
`dimethylsilyl group, followed by carbamylation of the
`31-position with an isocyanate with the general struc-
`ture shown above. Removal of the protecting group
`provides the 31-carbamylated compounds. In the case
`of the tert-butyl dimethylsilyl protecting group, depre-
`tection can be accomplished under mildly acidic condi-
`tions.
`.
`Having the 31-position carbamylated and the 42-posi-
`tion deprotected, the 42-position can be reacted with a
`different
`isocyanate than was reacted with the 31-
`alcohol, to give compounds having different carba-
`mates at the 31~ and 42-positions. Alternatively, the
`42-carbamylated compounds, prepared as described
`above, can be reacted with a different isocyanate to
`provide compounds having different carbamates at the
`31- and 42-positions.
`The isocyanates used to prepare the compounds of
`the invention are commercially available or can be pre-
`pared by methods that are disclosed in the literature.
`March [Advanced Organic Chemistry, 3d ed., pp. 391,
`452, and 479 (1985)] describes a general method for
`preparing arylsulfonyl isocyanates,
`that can be used
`
`Ex. 1 122-0002
`
`Ex. 1122-0002
`
`
`
`3250; or
`H -———9CXSO20H ArSOzOI-l
`
`2V
`ArSOzCl Etc—09 msocho
`
`Immunosuppressive activity was evaluated in an in
`vitro standard pharmacological test procedure to mea-
`sure lymphocyte proliferation (LAF) and in two in vivo
`standard pharmacological test procedures. The first in
`vivo procedure was a popliteal lymph node (PLN) test
`procedure which measured the effect of compounds of
`this invention on a mixed lymphocyte reaction and the
`second in vivo procedure evaluated the survival time of
`a pinch skin graft. The ability to prevent or treat trans-
`plantation rejection was also demonstrated in an in vivo
`heart allograft standard test procedure.
`The cornitogen-induced thymocyte proliferation pro-
`cedure (LAF) was used as an in vitro measure of the
`immunosuppressive effects of
`representative com-
`pounds. Briefly, cells from the thymus of normal
`BALB/c mice are cultured for 72 hours with PHA and
`IL-1 and pulsed with tritiated thymidine during the last
`six hours. Cells are cultured with and without various
`concentration of rapamycin, cyclosporin A, or test
`compound. Cells are harvested and incorporated radio-
`activity is determined. Inhibition of lymphoprolifera—
`tion is assessed as percent change in counts per minute
`from non-drug treated controls. The results are ex-
`pressed according to the following ratio.
`
`JH-control thmus cells — H3-rapamycin-treated thymus cells
`3I'I-control thymus cells — Ila-test compound-treated cells
`
`A mixed lymphocyte reaction (MLR) occurs when
`lymphoid cells from genetically distinct animals are
`combined in tissue culture. Each stimulates the other to
`undergo blast transformation which results in increased
`DNA synthesis that can be quantified by the incorpora-
`tion of tritiated thymidine. Since stimulating a MLR is a
`function of disparity at Major Histocompatibility anti-
`gens, an in vivo popliteal lymph node (PLN) test prooe~
`dure closely correlates to host vs. graft disease. Briefly,
`irradiated spleen cells from BALB/c donors are in-
`jected into the right hind foot pad of recipient C3H
`mice. The drug is given daily, p.o. from Day 0 to Day
`4. On Day 3 and Day 4, tritiated thymidine is given i.p.,
`b.i.d. On Day 5, the hind popliteal lymph nodes are
`removed and dissolved, and radioactivity counted. The
`corresponding left PLN serves as the control for the
`PLN from the injected hind foot. Percent suppression is
`calculated using the non-drug treated animals as allo-
`genic control. Rapamycin at a dose of 6 mg/kg, p.o.
`gave 86% suppression, whereas cyclosporin A at the
`same dose gave 43% suppression. Results are expressed
`by the following ratio:
`
`5
`
`15
`
`20
`
`25
`
`30
`
`35
`
`45
`
`55
`
`65
`
`5, 194,447
`
`3
`where the arylsulfonylisocyanate is not commercially
`available. The following scheme is illustrative of one
`method starting from an aryl moiety. Other methods of
`preparing arylsulfonyl isocyanates are known in the
`literature.
`
`4
`
`3
`
`3H-PLN cells control cm mouse —
`- N n
`'n-tr
`t
`H—PLN cells control CBH mouse —
`3H-PLN cells test compound-treated C311 mouse
`
`The second in vivo test procedure is designed to
`determine the survival time of pinch skin graft from
`male DBA/Z donors transplanted to male BALE/c
`recipients. The method is adapted from Billingham R.
`E. and Medawar P. B., J. Exp. Biol. 28:385-402, (1951).
`Briefly, a pinch skin graft from the donor is grafted on
`the dorsum of the recipient as a homograft, and an
`autograft is used as control in the same region. The
`recipients are treated with either varying concentra-
`tions of cyclosporin A as test control or the test com-
`pound, intraperitoneally. Untreated recipients serve as
`rejection control. The graft is monitored daily and ob-
`servations are recorded until the graft becomes dry and
`forms a blackened scab. This is considered as the rejec—
`tion day. The mean graft survival time (number of
`days-LSD.) of the drug treatment group is compared
`with the control group.
`The following table summarizes the results of repre-
`sentative compounds of this invention in these three
`standard test procedures.
`TABLE 1
`LAF
`PLN
`Skin Graft
`
`Compound
`(ratio)
`(ratio)
`(days i- SD)
`Example 1
`0.91
`0.84 (13.0.)
`11.2 :L- 0.4
`1.03 (i.p.)
`+
`0.47
`0.74
`Example 3
`+
`+
`0.10
`Example 4
`Rapamycin 12.0 i- 1.7 LCO 1.00
`
`
`
`+ Not evaluated.
`
`The results of these standard pharmacological test
`procedures demonstrate immunosuppressive activity
`both in vitro and in vivo for the compounds of this
`invention. Positive ratios in the LAF and PLN test
`procedures indicate suppression of T cell proliferation.
`As a transplanted pinch skin grafts are typically re-
`jected within 6-7 days without the use of an immuno-
`suppressive agent, the increased survival time of the
`skin graft when treated with the compounds of this
`invention further demonstrates their utility as immuno-
`suppressive agents.
`The ability of the compounds of this invention to
`prevent or treat transplantation was demonstrated in a
`heterotropic heart allograft standard pharmacological
`test procedure that emulates transplantation rejection
`that occurs in humans. The following briefly describes
`the procedure that was used. Male BN rat neonate do-
`nors (less than 5 days of age) were humanely sacrificed,
`the thymus was dissected away from the heart. All
`connections with the thoracic cavity were severed and
`the heart was removed from the chest cavity and placed
`in cooled RPMI media where all adherent fat and fascia
`were removed. The heart was bisected in half, along the
`midline from the apex to the root of the aorta, to gener-
`ate two approximately equal halves each containing
`atrial and ventricular tissue. Recipient male Lewis rats
`were anesthetized with phenobarbitol (50 mg/mL; i.p.),
`the left inner ear was swabbed with povidine iodine, and
`1 mL RPMI was injected subcutaneously above the
`cartiledge plate to produce a fluid filled sac. A stab
`incision was made to the sac, into which was inserted a
`single half heart fragment. The pocket was sealed with
`
`Ex. 1 122-0003
`
`Ex. 1122-0003
`
`
`
`5
`a single drop-of Vet-Seal (3M Animal Care Products).
`Recipients were divided into treatment groups of 10 rats
`each and the compounds to be evaluated were adminis-
`tered at a dosage of 225 lag/day following the transplan-
`tation procedure until graft failure occurred. Adminis-
`tration was i.p., either by manual injection or via an
`Azlet osmotic pump that was implanted into the perito-
`neum of the recipient rat. Grafts were inspected for loss
`of cardiac activity on day 7 post-transplant and subse-
`quently on alternate days. Graft survival time is defined
`as the post-transplant day on which the heart graft has
`lost all contractile activity by visual inspection and/or
`cardiac monitor. Individual rejection times were aver-
`aged to produce a mean survival time for each treated
`group. The following table shows the results obtained
`for a representative compound of this invention. The
`control group represents rats that received the allograft
`and no test compound. Rapamycin was evaluated for
`the purpose of comparison.
`20
`TABLE 2
`
`Mean Survival Time (days)
`Treatment Group
`11.5
`Control
`34.4
`Example 1
`38
`
`Rapamycin
`
`5
`
`10
`
`15
`
`25
`
`30
`
`35
`
`45
`
`As shown in Table 2, treatment with a representative
`compound ofthis inventiOn significantly (p<0.05) pro-
`longed survival time of the transplanted allograft. The
`untreated allograft was rejected by 11.5 days, whereas
`treatment with a representative compound of this in-
`vention prevented rejection until 34.4 days. As such, the
`results of this standard pharmacological test procedure
`demonstrate that the compounds of this invention are
`useful in preventing or treating transplantation rejec-
`tion.
`Because the compounds of this invention are structur-
`ally similar to rapamycin and have a similar activity
`profile to rapamycin, the compounds of this invention
`also are considered to have antitumor and antifungal
`activities.
`Based on the results of these standard pharmacologi-
`cal test procedures, the compounds are useful in the
`treatment of transplantation rejection such as, heart,
`kidney, liver, bone marrow, and skin transplants; auto-
`immune diseases such as, lupus, rheumatoid arthritis,
`diabetes mellitus, myasthenia gravis, and multiple scle-
`rosis; and diseases of inflammation such as, psoriasis,
`dermatitis, eczema, seborrhea,
`inflammatory bowel
`disease, and eye uveitis; solid tumors; and fungal infec»
`tions.
`As the compound of Example 3 was prepared via its
`42-si1y1ated intermediate (Example 2), the compound of
`Example 2 is therefore a useful as an intermediate of
`compounds of this invention.
`The compounds of this invention can be formulated 55
`neat or with a pharmaceutical carrier to a mammal in
`need thereof. The pharmaceutical carrier may be solid
`or liquid.
`A solid carrier can include one or more substances
`which may also act as flavoring agents, lubricants, solu-
`bilizers, suspending agents, fillers, glidants, compression
`aids, binders or tablet—disintegrating agents; it can also
`be an encapsulating material. In powders, the carrier is
`a finely divided solid which is in admixture with the
`finely divided active ingredient. In tablets, the active
`ingredient is mixed with a carrier having the necessary
`compression properties in suitable proportions and
`compacted in the shape and size desired. The powders
`
`'50
`
`65
`
`5, 194,447
`
`6
`and tablets preferably contain up to 99% of the active
`ingredient. Suitable solid carriers include, for example,
`calcium phosphate, magnesium stearate, talc, sugars,
`lactose, dextrin, starch, gelatin, cellulose, methyl cellu-
`lose, sodium carboxymethyl cellulose, polyvinylpyr-
`rolidine, low melting waxes and ion exchange resins.
`Liquid carriers are used in preparing solutions, sus-
`pensions, emulsions, syrups, elixirs and pressurized
`compositions. The active ingredient can be dissolved or
`suspended in a pharmaceutically acceptable liquid car-
`rier such as water, an organic solvent, a mixture of both
`or pharmaceutically acceptable oils or fats. The liquid
`carrier can contain other suitable pharmaceutical addi-
`tives such as solubilizers, emulsifiers, buffers, preserva-
`tives, sweeteners, flavoring agents, suspending agents,
`thickening agents, colors, viscosity regulators, stabiliz-
`ers or osmo-regulators. Suitable examples of liquid car-
`riers for oral and parenteral administration include
`water (partially containing additives as above, e.g. cel-
`lulose derivatives, preferably sodium carboxymethyl
`cellulose solution), alcohols (including monohydric
`alcohols and polyhydric alcohols, e.g. glycols) and their
`derivatives, and oils (e.g. fractionated coconut oil and
`arachis oil). For parenteral administration, the carrier
`can also be an oily ester such as ethyl oleate and isopro—
`pyl myristate. Sterile liquid carriers are useful in sterile
`liquid form compositions for parenteral administration.
`The liquid carrier for pressurized compositions can be
`halogenated hydrocarbon or other pharmaceutically
`acceptable propellent.
`Liquid pharmaceutical compositions which are sterile
`solutions or suspensions can be utilized by, for example,
`intramuscular,
`intraperitoneal or subcutaneous injec-
`tion. Sterile solutions can also be administered intrave-
`nously. The compound can also be administered orally
`either in liquid or solid composition form.
`The compounds of this invention may be adminis-
`tered rectally in the form of a conventional suppository.
`For administration by intranasal or intrabronchial inha-
`lation or insufflation, the compounds of this invention
`may be formulated into an aqueous or partially aqueous
`solution, which can then be utilized in the form of an
`aerosol. The compounds of this invention may also be
`administered transdermally through the use of a trans-
`dermal patch containing the active compound and a
`carrier that is inert to the active compound, is non toxic
`to the skin, and allows delivery of the agent for systemic
`absorption into the blood stream via the skin. The car—
`rier may take any number of forms such as creams and
`ointments, pastes, gels, and occlusive devices. The
`creams and ointments may be viscous liquid or semi—
`solid emulsions of either the oil-in-water or water-in-oil
`type. Pastes comprised of absorptive powders dispersed
`in petroleum or hydrophilic petroleum containing the
`active ingredient may also be suitable. A variety of
`occlusive devices may be used to release the active
`ingredient into the blood stream such as a semipermia-
`ble membrane covering a reservoir containing the ac-
`tive ingredient with or without a carrier, or a matrix
`containing the active ingredient. Other occlusive de-
`vices are known in the literature.
`Preferably, the pharmaceutical composition is in unit
`dosage form, e.g. as tablets or capsules. In such form,
`the composition is sub-divided in unit dose containing
`appropriate quantities of the active ingredient; the unit
`dosage forms can be packaged compositions, for exam-
`ple, packeted powders, vials, ampoules, prefilled syrin-
`
`Ex. 1 122-0004
`
`Ex. 1122-0004
`
`
`
`7
`ges or sachets containing liquids. The unit dosage form
`can be, for example, a capsule or tablet itself, or it can be
`the appropriate number of any such compositions in
`package form. The dosage to be used in the treatment
`must be subjectively determined by the attending physi-
`can.
`
`In addition, the compounds of this invention may be
`employed as a solution, cream, or lotion by formulation
`with pharmaceutically acceptable vehicles containing
`0.1-5 percent, preferably 2%, of active compound
`which may be administered to a fungally affected area.
`The following examples illustrate the preparation of
`representative compounds of this invention.
`EXAMPLE 1
`
`Rapamycin 42-ester with phenylsulfonylcarbamic acid
`A solution of 192 mg of rapamycin in 1 mL toluene
`was treated at 0' under nitrogen with 37 mg of benzene-
`sulfonyl isocyanate in 1 mL toluene. After stirring at 0'
`under nitrogen for one hour, the reaction mixture was
`diluted with 100 ml of ethyl acetate and washed with
`brine (3X20 ml). The ethyl acetate solution was dried
`with magnesium sulfate and evaporated. The residue
`was chromatographed on silica gel. Elation with ethyl
`acetate/hexane (1:1) afforded 36 mg of the title com-
`pound as a white foam.
`IR (KBr) 3440 (OH), 2910, 1735 & 1710 (C=O), 1635
`(aromatic), 1440, 1160, 1080 and 980 cm—1. NMR
`(CDC13, 400 MHz): 5 8.04 (m, 2H, aromatic), 7.65 (m,
`1H, aromatic), 7.56 (m, 2H, aromatic), 3.41 (s, 3H,
`OCH3), 3.34 (s, 3H, 0CH3), 3.13 (s, 3H, OCH3). MS
`(neg. ion FAB): 1095 (M—), 590. 321.
`EXAMPLE 2
`
`Rapamycin 42-tertbutyldimethylsi1yl ether-31-ester
`with phenyl sulfonyl carbamic acid
`Raparnycin (0.914 g) was added to a solution of 0.204
`g imidazole and 0.165 g tert-butyldimethylsilyl chloride
`in 4 mL dimethylformamide at 0° and stirred under
`nitrogen for 16 hours at 20'. Brine (100 mL) was added,
`the product was extracted into ether, washed with
`brine, dried over MgSO4 and evaporated. Chromatog-
`raphy on silica gel afforded 0.65 g of rapamycin-42-tert-
`butyldimethylsilyl ether as a white solid.
`A solution of 308 mg of rapamycin-42-tert-butyldime-
`thylsilyl ether in 1 mL toluene was treated at 0' under
`nitrogen with 109 mg of benezenesulfonyl isocyanate in
`1 mL toluene. After stirring at 0‘ under nitrogen for
`three hours, the reaction mixture was diluted with 100
`mL ethyl acetate and washed with brine 2 X20 ml). The
`ethyl acetate solution was dried with magnesium sulfate
`and evaporated. The residue was chromatographed on
`silica gel. Elution with ethyl acetate/hexane (1:1) af-
`forded 310 mg of the title compound as a white foam.
`IR (KBr): 3400 and 3240 (OH), 2915, 1740 and 1710
`(C=O), 1640, 1440, 1160, 1080 cm-1. NMR (CDC13,
`400 MHz): 6 8.00 (m, 2H, aromatic), 7.55 (m, 3H, aro-
`matic), 3.45 (s, 3H, OCH3), 3.27 (s, 3H, OCH3), 3.13 (s,
`3H, OCH3). MS (neg. ion FAB): 1209 (M-), 590, 156.
`EXAMPLE 3
`
`Rapamycin-3l-ester with phenylsulfonylcarbamic acid
`A solution of 250 mg of Rapamycin-42-tert-butyl-
`dimethylsilyl ether-31-ester with phenylsulfonylcar-
`bamic acid in 2 mL of tetrahydrofuran was treated with
`2 mL water and 6 mL acetic acid. After stirring at room
`temperature under nitrogen for 20 hours, the mixture
`
`5, 194,447
`
`was diluted with 130 mL ethyl acetate and washed with
`brine (2x20 ml). The ethyl acetate solution was dried
`with magnesium sulfate and evaporated to dryness to
`give 208 mg of the title compound as a white foam.
`IR (KBr): 3400 (OH), 2930, 1730 (C=O), 1640 (aro-
`matic), 1440, 1340, 1160, 1085 cm—1. NMR (CDC13, 400
`MHz): 6 8.0 (m, 2H, aromatic), 7.59 (m, 3H, aromatic),
`3.41 (s, 3H, OCH3), 3.26 (s, 3H, OCH3). MS (neg. ion
`FAB): 1095 (M‘), 589, 321.
`EXAMPLE 4
`
`V Rapamycin 42-ester with
`(4—chlorophenylsulfonyl)carbamic acid
`A solution of 400 mg of rapamycin in 10 mL ether/-
`toluene (1:4) was treated at — 10' under nitrogen with
`100 mg of 4-chlorobenzenesulfouylisocyanate in 1 mL
`toluene. The mixture was stirred at 0' under nitrogen
`for 3.5 hours, diluted with 120 mL of ethyl acetate,
`washed with saturated sodium bicarbonate solution,
`followed by a water wash, and dried with magnesium
`sulfate. The ethyl acetate solution was evaporated and
`the residue chromatographed on silica gel. Elution with
`ethyl acetate afforded 100 mg of the title compound as
`a white solid, mp 142'—146’. The title compound was
`isolated as a trihydrate.
`1H NMR (CDC13, 400 MHz), 6 8.0 (d, J: 13 cps, 2H,
`aromatic protons meta to sulfonyl group), 7.53 (d, J = 13
`cps, 2H, aromatic protons ortho to sulfonyl group), 3.30
`(s, 3H, 0CH3), 3.20 (s, 3H, OCH3), 3.13 (s, 3H, OCH3)
`ppm. MS (neg ion FAB): 1129 (MH-). IR KBr max 3430
`(OH), 2930, 1740, 1730 (both C20), 1640 (amide
`C=O), 1450, 1350 (—-S02—). 1160 (—SOz—), 1160
`(—SOZ—-), 1090, 985 cm—1.
`Anal. Calcd. for C53H33CIN201553 H20: C 58.74; H
`7.56; N 2.35 Found: C 58.92; H 7.99; N 2.17
`
`EXAMPLE 5
`Rapamycin 42-ester with (3-methylphenylsulfonyl)
`carbamic acid
`
`A solution of400 mg rapamycin in 10 mL ethyl aceta-
`te/ether (k) was treated at — 10‘ under nitrogen with
`127 mg of 3-methylphenylsulfonyl isocyanate in 2 mL
`ether. THe mixture was stirred at 0' under nitrogen for
`4.5 hours, diluted with 120 mL ethyl acetate, washed
`with a saturated sodium bicarbonate solution, and dried
`with magnesium sulfate. The ethyl acetate solution was
`evaporated and the residue chromatographed on silica
`gel. Elution with ethyl acetate/n-hexane (1:1) afforded
`270 mg of the title compound as a white powder, mp
`116‘—120'.
`1H NMR (CDC13, 400 MHz) 5 8.13 (d, J=13 cps, 1H,
`aromatic proton ortho to sulfonyl group), 7.53 (m, 1H,
`aromatic proton, para to sulfonyl group), 7.39 (m, 1H,
`aromatic protons para to methyl group), 7.33 (d, J = 13
`cps, 1H, aromatic ortho to methyl group). 3.23 (s, 3H,
`OCH3), 3.18 (s, 3H, OCHS), 3.14 (s, 3H, 0CH3), 2.67 (s,
`3H, aromatic methyl) ppm. MS (neg. ion FAB): 1109
`(MH- ). IR KBr max 3450 (OH), 2940, 1740, 1720
`(C=O), 1640 (amide C=0 , 1450, 1340 (——SOz—),
`1160 (—SOz—), 990 cm-1.
`EXAMPLE 6
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`Rapamycin 31,42-diester with phenylsulfonylcarbamic
`acid
`
`A solution of 200 mg of rapamycin 42-ester with
`phenylsulfonylcarbamic acid in 3 mL ethyl aceta-
`
`Ex. 1 122-0005
`
`Ex. 1122-0005
`
`
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`9
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`5, 194,447
`
`
`
`wherein
`independently, hydrogen or
`R1 and R2 are each,
`—CONHSOz-Ar; and Ar is phenyl, naphthyl, pyri-
`dyl, quinolyl,
`isoquinolyl, quinoxalyl,
`thienyl,
`thionaphthyl, fury], benzofuryl, benzodioxyl, benz-
`oxazolyl,
`benzoisoxazolyl,
`or
`benzodioxolyl;
`wherein the Ar group may be optionally mono-,
`di-, or tri-substituted with a group selected from
`alkyl of 1—6 carbon atoms, phenylalkyl of 7—10
`carbon atoms, alkoxy of 1—6 carbon atoms, cyano,
`halo, nitro, carbalkoxy of 2—7 carbon atoms, tn'flu-
`oromethyl, amino, dialkylamino of 1—6 carbon
`atoms per alkyl group, alkylthio of 1-6 carbon
`atoms, —SO3H, —P03H, and -—C02H;
`with the proviso that R1 and R2 are not both hydrogen;
`or a pharmaceutically acceptable salt thereof when the
`Ar group contains a basic nitrogen or when the Ar
`group is substituted by dialldyamino of 1-6 carbon
`atoms per alkyl group, —S()3H, ——PO3H, or —C02H.
`2. A compound of claim 1 wherein Ar is phenyl; the
`phenyl group being optionally mono-, di-, or tri-sub-
`stituted or a pharmaceutically acceptible salt thereof.
`3. A compound of claim 1 which is rapamycin 42-
`ester with phenylsulfonylcarbamic acid.
`4. A compound of claim 1 which is rapamycin-31-
`ester with phenylsulfonylcarbamic acid.
`5. A compound of claim 1 which is rapamycin 42-
`ester with (4-chloropheny1su1fony1)carbamic acid.
`6. A compound of claim 1 which is rapamycin 42-
`ester with (3-methylphenylsu1fonyl) carbamic acid.
`7. A compound which is rapamycin 42-tertbutyldime-
`thylsilyl ether-Slcster with phenylsulfonylcarbamic
`acid.
`.
`l
`t
`.
`I
`
`te/ether (1:1) was treated at 0‘ under nitrogen with 200
`
`mg of benzenesulfonyl isocyanate in 0.5 mL of ether.
`
`After stirring at 0' under N2 for 18 hours and at 22'
`
`under N2 for 3 hours, the reaction mixture was diluted
`
`with 120 mL ethyl acetate, washed with a saturated
`
`sodium bicarbonate solution, and then washed with
`
`brine. The ethyl acetate solution was dried with magne-
`
`sium sulfate and evaporate. The residue was chromato-
`
`graphed on silica gel. Elution with ethyl acetate af-
`
`forded 75 mg of the title compound as a white foam, mp
`
`104'-108°.
`
`NMR (CDClg, 400 MHz): ‘0‘ 8.03 (m, 2H, aromatic),
`
`7.55 (m, 3H, aromatic), 3.26 (s, 3H, OCH3), 3.22 (s, 3H,
`
`OCH3), 3.13 (s, 3H, OCH3) ppm. MS (neg. ion FAB):
`
`1278 (MH—). IR (KBr): 3340 (OH), 2930, 1730 & 1715
`
`((1:0), 1630 (aromatic), 1445, 1290, 1160, 1095, 990
`
`cm — 1.
`
`What is claimed is:
`
`l. A compound of the formula
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`Ex. 1 122-0006
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`Ex. 1122-0006
`
`